This project has published evidence that the Rho(D) antigenic determinants are associated with the major membrane polypeptide of the red cell, band 3. The proposed research seeks to identify the membrane polypeptides bearing the other major antigens in the Rh blood group, C, c, E and e. The use of reversible crosslinking reagents will alsow immunospecific reversible attachment of antibody to native antigenic determinants in intact red cells. Following enzymatic or chemical degradation and the appropriate use of multiple radioisotopes, techniques of peptide analysis should provide direct information about the amino acid residues comprising Rh antigenic determinants. Antigen-antibody complexes solubilized in mild detergent will be characterized by hydrodynamic methods designed to provide information about size, shape and stoichiometry of the complexes. A working hypothesis to be tested postulates that all of the Rh antigens may represent different oligomeric forms of band 3. Experiments involving the use of radiolabeled sulfhydryl reagents and peptide analysis may reveal patterns associated with each of the Rh antigens if band 3 oligomeric associations are dependent on sulfhydryl oxidation-reduction. The Rh blood group is influenced reversibly both by phospholipids and cholesterol. The project will examine the stuctural or physicochemical features responsible for lipid modulation of all the Rh antigens. The Rh complex is important to red cell survival and to our understanding of some autoimmune hemolytic anemias. The elucidation of the molecular biology of Rh will further our understanding of this clinically important blood group and of biological membranes in general.